靶向性CPI-444载药纳米微粒的制备及其对T细胞活性和抗肿瘤效应的影响OA北大核心CSTPCD

Objective:To prepare and characterize CD8 antibody-conjugated CPI-444(C)-loaded nanoparticles(CNP/α CD8)and investigate their effects on CD8+T cell activation,proliferation,and anti-tumor activity.Methods:Nanoparticles encapsulating the adenosine A2A receptor(A2AR)antagonist CPI-444(C)or the fluorescent dye Coumarin-6(C6)were prepared using the double emulsion solvent evaporation method and EDC/NHS chemistry for antibody conjugation,resulting in CNP/αCD8 and C6NP/αCD8.The morphology and size of the nanoparticles were characterized by scanning electron microscopy and NanoPlus particle size analyzer.Drug loading and release profiles were determined using liquid chromatography-tandem mass spectrometry(LC-MS/MS)and centrifugation.The internalization of C6NP/αCD8 by CD8+T cells were examined by flow cytometry and fluorescence microscopy.The effects of CNP/αCD8 on the proliferation,activation,cytotoxicity,and tumor killing ability of CD8+T cells were examined by flow cytometry,ELISA,and lactate dehydrogenase(LDH)assay.Results:The CNP/αCD8 nanoparticles were spherical with an average diameter of about 150 nm,effectively encapsulating CPI-444 and conjugating CD8 antibodies,with a drug encapsulation efficiency and a CD8 antibody conjugation efficiency of approximately 60%and 53.4%,respectively.The nanoparticles exhibited good stability and were efficiently internalized by CD8+T cells,inhibiting A2AR expression.Biological function assays showed that CNP/αCD8 enhanced CD8+T cell proliferation,promoted T cell activation,cytokine secretion,granzyme B,and perforin production,and improved the tumor-killing ability of CD8+T cells.Conclusion:CNP/αCD8 nanoparticles can significantly enhance the immune functions of CD8+T cells,likely by inhibiting A2AR expression.